Selective detection of alkenes or alkynes

ABSTRACT

A detector can detect an analyte including a carbon-carbon multiple bond moiety and capable of undergoing Diels-Alder reaction with a heteroaromatic compound having an extrudable group. The detector can detect, differentiate, and quantify ethylene.

PRIORITY CLAIM

This application is a divisional of U.S. patent application Ser. No.14/246,008, filed Apr. 4, 2014, now U.S. Pat. No. 10,545,093, whichclaims priority to U.S. Provisional Patent Application No. 61/809,362,filed Apr. 6, 2013, which is incorporated by reference in its entirety.

STATEMENT OF GOVERNMENT SUPPORT

This invention was made with government support under Contract No.W911NF-07-D-0004 awarded by the Army Research Office. The government hascertain rights in the invention.

TECHNICAL FIELD

This invention relates to detection methods and detectors for sensingalkenes or alkynes.

BACKGROUND

The detection and monitoring of alkene and alkyne is of great interestand importance. Current methods for alkene and alkyne detection,differentiation and quantitation are expensive and there is a growingneed of a method that is low-cost and ease-of-use. Containingunsaturated functional groups, alkene and alkyne are capable ofundergoing Diels-Alder type reactions.

SUMMARY

In one aspect, a detector for detecting an analyte can include a housingincluding a detection region comprising a compound having an extrudablegroup and capable of undergoing Diels-Alder reaction with the analyteincluding a carbon-carbon multiple bond moiety. The compound can be aheteroaromatic compound.

In certain embodiments, the detector can be a color based detector. Inother embodiments, the detector can be a fluorescence based detector. Inother embodiments, the detector can be a resistivity based detector.

In some embodiments, the analyte can include ethylene. In someembodiments, the heteroaromatic compound can include tetrazine. In someembodiments, the heteroaromatic compound can includebis-2-pyridyl-1,2,4,5-tetrazine.

In some embodiments, the heteroaromatic compound can selectively reactwith a sterically unencumbered alkene. In some embodiments, theheteroaromatic compound can selectively react with a stericallyunencumbered alkyne.

In some embodiments, the detection region can include a colorimetricindicator that changes color after the Diels-Alder reaction. In someembodiments, the color based detector can include a copper salt, anickel salt, a silver salt, a zinc salt, an aluminum salt, or a goldsalt. In some embodiments, the detection region can include a machinereadable pattern. In some embodiments, the color based detector caninclude a reader capable of reading the pattern.

In some embodiments, the detection region can include other materialsthat report on the state of the molecule, or collection of molecules,that undergo Diels-Alder reactions. For example, the detection regioncan include a material initially having low luminescence and after theDiels-Alder reaction with an analyte the composition will have anincreased luminescence. In other examples, the detection region caninclude a material initially having high luminescence and after theDiels-Alder reaction with an analyte the composition will have reducedluminescence. Multiple materials with different luminescent propertiescan be used to provide for light emission and absorbance patterns thatcan be read. In these embodiments, the detection region can include aluminescent material. Examples of suitable luminescent materials caninclude one or more of a conjugated molecule, a conjugated polymer, oran inorganic phosphor.

In other embodiments, the detection region can include an electricallyconductive material and the measured response can be a change in theresistivity in the detection region. In some cases, the Diels-Alderreaction with an analyte can increase the resistance in the detectionregion. In other cases, the Diels-Alder reaction with an analyte candecrease the resistance in the detection region. In these embodiments,the detection region can include one or more of a carbon nanotube,graphite, graphene, a semiconductor nanowire, a metal nanoparticle, ametal nanowire, or a conducting polymer.

In another aspect, a method of detecting an analyte in a sampleincluding a carbon-carbon multiple bond moiety can comprising exposing adetection region of a detector including a compound having an extrudablegroup and capable of undergoing Diels-Alder reaction with the analyteincluding a carbon-carbon multiple bond moiety to the sample, anddetecting a property change of a reaction mixture comprising theheteroaromatic compound based on the presence of the analyte in thesample. In some embodiments, the analyte can be ethylene.

In some embodiments, detecting the property change can includemonitoring absorbance of the detection region. In some embodiments,detecting the property change can include monitoring fluorescence of thedetection region. In some embodiments, detecting the property change caninclude monitoring conductivity of the detection region. In someembodiments, the method can further include determining a reaction rateconstant of the analyte with the compound in the detection region of thedetector. In some embodiments, the method can further includequantifying the amount of the analyte using the rate constant.

In some embodiments, the method can further include differentiatingalkene classes, alkyne classes, or alkene and alkyne classes bycorrelating reactivity with initial and final colors of the reactionmixture.

In some embodiments, the method can further include quantifying theamount of the analyte using Red Green Blue color mapping. In someembodiments, the method can further include differentiating alkeneclasses, alkyne classes, or alkene and alkyne classes using Red GreenBlue color space value. In some embodiments, the method can furtherinclude quantifying the amount of the analyte using Euclidean distancedetermined from the initial and final colors of the reaction mixture. Insome embodiments, the method can further include differentiating alkeneclasses, alkyne classes, or alkene and alkyne classes using euclideandistance.

In some embodiments, the method can further include reading amachine-readable pattern in the detection region when the patternappears. In some embodiments, the method can further include reading amachine-readable pattern in the detection region when the patternvanishes.

In some embodiments, the reaction mixture can be in a solvent. In someembodiments, the solvent can include tetrahydrofuran, dichloromethane,acetonitrile, nitromethane, toluene, chloroform, propylene carbonate,dimethylsulfoxide, dimethylformamide or acetone. In some embodiments,the reaction mixture can be in solid formulation or solid substrate. Insome embodiments, the reaction mixture can include a copper salt, anickel salt, a silver salt, a zinc salt, an aluminum salt, or a goldsalt.

In another aspect, a detector for detecting an analyte can include ahousing including a detection region comprising a compound havingelectron donating or withdrawing character and capable of undergoingDiels-Alder reaction with the analyte including a carbon-carbon multiplebond moiety, and a conducting material that changes conductivity afterthe Diels-Alder reaction.

In certain embodiments, the compound can become less electron acceptingafter reacting with an alkene. The compound can become less electrondonating after reacting with an alkene. Resistivity of the conductingmaterial can change after the Diels-Alder reaction. The compound canselectively react with a sterically unencumbered alkene.

In certain embodiments, the detection region can include a colorimetricindicator that changes color after the Diels-Alder reaction. Thedetector can include a copper salt, a nickel salt, a silver salt, a zincsalt, an aluminum salt, or a gold salt. The detection region can includea machine readable pattern. The detector can include a reader capable ofreading the pattern.

In certain embodiments, the detector can be a color based detector. Thedetector can be a fluorescence based detector. The detector can beresistivity based detector.

In certain embodiments, the detector can selectively detect anelectron-poor alkene. The detector can selectively detect anelectron-rich alkene. The selectivity for an electron-poor alkene can bewith respect to an electron-rich alkene. The selectivity for anelectron-rich alkene can be with respect to an electron-poor alkene.

In another aspect, a method of detecting an analyte in a sampleincluding a carbon-carbon multiple bond moiety can include exposing adetection region of a detector including a compound having electrondonating or withdrawing character and capable of undergoing Diels-Alderreaction with the analyte including a carbon-carbon multiple bondmoiety, and detecting a property change of a conducting material afterthe Diels-Alder reaction based on the presence of the analyte in thesample.

Other aspects, embodiments, and features will be apparent from thefollowing description, the drawings, and the claims.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a block diagram showing one example of a detecting device.

FIG. 2 is a demonstration of selective alkene detection.

FIG. 3A shows time based color change when the solution contains silvertriflate and bptz. FIG. 3B shows one reaction example.

FIG. 4 shows the extent of color change post-ethylene addition when thesolution contains different kinds of silver salt.

FIG. 5 shows photographs of vials containingbis-2-pyridyl-1,2,4,5-tetrazine (bptz) and additive in dichloromethane,before exposure to ethylene and after 1.5 h of exposure to a headspaceof C₂H₄ (1 atm) at room temperature.

FIG. 6 shows euclidean distance as a quantitative representation ofreaction color profile.

FIG. 7 shows a representative example of a UV-VIS spectrum of Bptz inorganic solvent.

FIG. 8 shows graphical representation of UV-VIS spectroscopicmeasurements of max (λ_(max)=545 nm) of a solution of Bptz inacetonitrile (MeCN) when exposed to a gross molar excess of C₂H₄ (1 atm)over time.

FIG. 9 shows kinetics experiment results in nitromethane (MeNO₂),dimethylformamide (DMF), and acetonitrile (MeCN).

FIG. 10 shows photographs of alkene sensing material incorporated into aglass tube containing glass wool.

FIG. 11 (panel A) shows a photograph of QR code printed in pink on copypaper; FIG. 11 (panel B) shows a photograph of Bptz impregnatedpolystyrene coating covering a printed Quick Read (QR) code on copypaper before exposure to C₂H₄; FIG. 11 (panel C) shows a photograph ofBptz impregnated polystyrene coating covering a printed QR code on copypaper after exposure to C₂H₄ (1 atm) for 1 h.

FIG. 12 (panel A) shows a photograph of UPC code; FIG. 12 (panel B)shows a photograph of Quick Read (QR) code; FIG. 12 (panel C) shows aphotograph of USPC data matrix; FIG. 12 (panel D) shows a photograph ofGS1 DataBar; FIG. 12 (panel E) shows a photograph depicting Bptz ink(pink colored solution indicated by arrow) with an example of a GS1DataBar that it was used to print on copy paper.

FIG. 13 (panel A-1) and FIG. 13 (panel B-1) shows photographs ofstandardized, machine-readable representations of data printed usinginks created with Bptz as pigment before exposure to C₂H₄; FIG. 13(panel B-1) and FIG. 13 (panel B-1) shows photographs of therepresentations after exposure to C₂H₄ (1 atm) for 48 h.

FIG. 14 is a UV-VIS kinetics plot showing change in absorbance over timeafter exposure of a solution of Bptz in the presence of two differentmolar equivalents of trifluoroacetic acid to ethylene gas, monitoring asingle wavelength at 410 nm.

FIG. 15 is a graph depicting a time-course experiment showing the changein the UV-VIS spectra of a solution of 1:1 Bptz:AgOCOCF₃ in MeNO₂ afterexposure to 1-hexene. 0.1 minute increments.

FIG. 16 shows photographs of a graph depicting test results of exemplarysensors.

FIG. 17 shows photographs of a demonstration of selective electron-richalkene detection.

FIG. 18 shows photographs of a demonstration of selective electron-pooralkene detection.

FIG. 19 shows photographs of a demonstration of selective electron-pooralkene detection.

FIG. 20 shows photographs of a demonstration of other tetrazinesselectively reaction with electron rich alkenes.

FIG. 21 shows photographs of a demonstration of complementary reactivityof Bptz and 1,3-diphenylisobenzofuran.

DETAILED DESCRIPTION

Compounds containing unsaturated functional groups are capable ofundergoing [4+2] cycloadditions (Diels-Alder type reactions), resultingin the loss of an extrudable group, such as dinitrogen. This structuralchange in the compound results in a change in its electronic propertiesof material including the compound, which are then transduced intovarious observable outputs, such as change in color, absorbance,fluorescence, or conductivity. These observable outputs allow for thedetermination of total amount of analyte exposure to device at varioustime points following initial exposure.

Diels-Alder Reaction

The Diels-Alder reaction is a cycloaddition reaction between aconjugated diene and a substituted alkene to form a substitutedcyclohexene system. Some of the atoms in the newly formed ring do nothave to be carbon. Some of the Diels-Alder reactions are reversible; thedecomposition reaction of the cyclic system is then called theretro-Diels-Alder.

The simplest example is the reaction of 1,3-butadiene with ethene toform cyclohexene:

The analogous reaction of 1,3-butadiene with ethyne to form1,4-cyclohexadiene is also known.

A heteroaromatic compound can be used for a Diels-Alder reaction. Theheteroaromatic compound can have an extrudable group as shown in thefollowing structure.

Each of A, B, Y and Z can be C—R, N, S, or O, such that Y and Z togetherform an extrudable group. Each R, independently, can be H, halo, alkyl,haloalkyl, or aryl.

An extrudable group is a group that leaves the heteroaromatic compoundafter the heteroaromatic compound undergoes a Diels-Alder reaction. Oneexample of the extrudable group is di-nitrogen. Examples of compoundswherein di-nitrogen is extrudable group:

The following is a schematic representation of inverse-electron-demandhetero-Diels-Alder reaction of (1) with an unsaturated carbon-carbonbond to give (2) via transition state (T.S. 1) (2) spontaneouslyundergoes a retro-Diels-Alder reaction resulting in product (3) byextrusion of (4).

Table 1 and Table 2 show examples of tetrazines.

TABLE 1

 

      isolated yield

a

commercial 535 415 19.41 b

25.4  540 very slow c

N/R d

18.4  535 425 6.9 e

22%  542 very slow

TABLE 2 f

2.4% 540 767 g

 48% 537 h

5.9% 534 417 i

0.5%  

R²     isolated yield

j

0.8% 547

Other tetrazines include bptz and tz-2:

Additional dienes capable of Diels Alder reactions for sensing ofalkenes can include structures that are highly electron deficient withelectron accepting properties including the following structures. Thesematerials additionally have extended aromatic structures that canpromote favorable interactions with the conducting material that theyare in contact with. After reaction with alkenes these materials can beless electron poor and the change in their electronic properties canserve to change the resistivity of the conducting materials. Theconducting materials can be based on conducting polymers, carbonnanotubes, graphene, graphite, metal nanowires, metal oxides, orinorganic semiconductors.

As a result of their electron poor nature these molecules can be mosteffective in Diels-Alder reactions with alkenes that are substitutedwith electron neutral or donating groups. These alkene substituents canbe other alkenes, alkyl groups, phenyl groups, or alkoxy groups. Thesemolecules can react with cyclic alkenes including highly strainedsystems including 1-methyl cycloproprene.

Multiple R groups can be added to the aromatic rings and the different Ror Z groups can be present in the same molecule. The aromatic can be aC6-C14 aryl, for example, phenyl or naphthyl. The alkyl can be a C1-C16alkyl, for example, methyl, ethyl, propyl, butyl, pentyl or hexyl. Thefluoroalkyl can be a fluorianted C1-C16 alkyl, for example, methyl,ethyl, propyl, butyl, pentyl or hexyl including 1, 2, 3, 4 or 5 fluorogroups.

A complement to the electron-poor molecules shown are electron-richDiels-Alder dienes that are shown below. These molecules are notexpected to undergo Diels-Alder reactions with ethylene and otherelectron rich alkenes. They are likely to be reactive with strainedalkenes or electron deficient alkenes. Of particular interest arealkenes of the formula CH₂CH(EWG), wherein EWG stands for an electronwithdrawing group such as a ester, ketone, nitrile, or carboxylic acid.Electron deficient alkenes such as acrylates and acrylonitrile are toxicchemicals that can be used in the polymer and finishes industries. Thesedienes can have high reactivity with these analytes. Upon reaction, theelectron donating properties of the diene can change and effect changesin the resistivity of a conducting materials in which that are incontact. The conducting materials can be based on conducting polymers,carbon nanotubes, graphene, graphite, metal nanowires, metal oxides, orinorganic semiconductors.

In both of the last sets or Diels-Alder dienes the reaction can cause alarge change in the electronic structure of the diene. This change cancause a change in the optical properties of the molecules and can causea change in the visible color of the compounds or changes in theirfluorescence intensity. These changes can be used to detect the presenceof an alkene analyte of interest.

It can be important to differentiate between electron poor DA dienes,often referred to as inverse demand DA dienes, and electron rich dienes.The inverse demand systems can be better with electron neutral ordonating alkenes, whereas the electron rich systems can be better withelectron poor alkenes which can be toxic.

Importance of Alkene and Ethylene Detection

Alkenes are an important feedstock, product, by-product, or transientspecies of many biological, laboratory scale, consumer, and industrialprocesses such as, but not limited to: biochemical pathways, internalcombustion engine exhaust, industrial manufacturing processes includingpetrochemical refining, oil & gas discovery, extraction, polymermanufacture, etc. The ability to selectively detect and quantify theirpresence is of interest in enabling a wide variety of applications, forexample, fruit ripeness determination, fruit & flower expiration dateprediction, breath analysis, oil & gas well characterization, industrialsafety, consumer safety at gas stations, exhaust characterization, andso on.

The detection and monitoring of ethylene is of great interest andimportance in the food and agricultural industries. Ethylene as one ofthe smallest plant hormones is responsible for the ripening of fruit andplays an important role in many more developmental plant processes suchas seed germination, fruit ripening, senescence and abscission.Measurement of the rate of ethylene evolution can be used as anindicator of fruit age. As fruits and vegetables start ripening,ethylene is produced and emitted, and the internal ethyleneconcentration in some fruits is used as a maturity index to determinethe time of harvest. As ripening begins, the production of ethylene canincrease dramatically. In some fruits and vegetables, such as bananas,the ripening process is continued after harvesting by exposure toethylene gas, and the monitoring of ethylene gas in storage rooms isimportant to avoid deterioration of ethylene-sensitive produce.

Methods of Alkene and Ethylene Detection

Current methods for alkene differentiation and quantitation exist butare either prohibitively expensive for many applications of interest(GC-FID-MS), prohibitively complex and/or unsafe for on-site use(titration with oxidizing agents such as KMnO₄, Br₂), or not selectiveenough for sophisticated applications beyond determining whether analkene is present (disposable alkene indicator tubes (Drager tubes),KMnO₄, Br₂).

Ethylene can be detected by different methods. A sensory system ofethylene can use fluorescent conjugated polymers. Esser, Birgit, andTimothy M Swager. “Detection of Ethylene Gas by Fluorescence Turn-On ofa Conjugated Polymer.” Angewandte Chemie International Edition 49.47(2010): 8872-8875, which is incorporated by reference its entirety. Thefluorescence of the conjugated polymer is partly quenched by thepresence of copper (I) moieties that can coordinate to the polymer. Uponexposure to ethylene gas, the copper complexes bind to the ethylenemolecules and no longer quench the polymer fluorescence. It requires aspecific binding event to the copper to create a new signal whereas afluorescence quench can take place. A chemoresistive sensor able todetect ethylene can be based on carbon nanotubes. Esser, Birgit, Jan MSchnorr, and Timothy M Swager. “Selective Detection of Ethylene GasUsing Carbon Nanotube-based Devices: Utility in Determination of FruitRipeness.” Angewandte Chemie International Edition 51.23 (2012):5752-5756, which is incorporated by reference in its entirety. Theethylene sensitive material is a mixture of SWNTs with a copper(I)complex 1 based upon a fluorinated tris(pyrazolyl) borate ligand, whichis able to interact with the surface of carbon nanotubes, therebyinfluencing their conductivity. Upon exposure to ethylene, complex 1binds to ethylene and forms complex 2, which has a decreased interactionwith the SWNT surface.

Methods of ethylene detection include Gas chromatography,High-performance liquid chromatography, nuclear magnetic resonance,drager tubes, combustion analysis and titration methods. But thesemethods are either expensive, or provide low-quality information.

Another method of detecting an analyte in a sample include acarbon-carbon multiple bond moiety comprising exposing a detectionregion of a detector including a heteroaromatic compound having anextrudable group and capable of undergoing Diels-Alder reaction with theanalyte including a carbon-carbon multiple bond moiety to the sample,and detecting color change of a reaction mixture comprising theheteroaromatic compound based on the presence of the analyte in thesample. This method provides alkene and alkyne detection,differentiation, and quantitation that addresses the growing need oftransducing relevant information (only previously attainable fromsophisticated methods such as GC-analysis) with the favorable low-costand ease-of-use attributes ascribed to more basic technologies. Usingthis method, a device can indicate the presence of specific classes ofalkenes or alkynes in the gas phase, and can determine the totalexposure of the device to said alkenes or alkynes, based on acolorimetric readout. Because this device is selective for certainclasses of alkenes and alkynes, it allows for differentiation ofcompounds of interest that contain certain alkene or alkynefunctionality. This method can make use of the color change thataccompanies the transformation of an s-tetrazine moiety to a pyrimidinemoiety upon reaction with unsaturated carbon-carbon bonds.

This approach to alkene detection has a number of useful characteristicsthat make it desirable as a method of alkene quantitation anddifferentiation. Some of these advantages include, but are not limitedto determining total exposure of an object to an alkene that mayinfluence it's properties, health, or former, current, or future stateof being and determining the presence of different classes of alkeneswith easy to understand, inexpensive technology. In addition, when theextrudable group of the heteroaromatic compound is di-nitrogen, theby-product of the reaction with alkyne or alkene, such as ethylene, isN₂, which is food safe.

Property Change after Diels-Alder Reaction

The properties of a detection region can change after the Diels-Alderreaction takes place. The property change can be a color change,fluorescence change, conductivity change or a combination thereof.

The extent of property change before and after Diels-Alder reactions canvary with different additives. Different colors can be obtained byadding a copper salt, a nickel salt, a silver salt, a zinc salt, analuminum salt, or a gold salt to the reaction mixture. Examples of theadditive include silver triflate (AgOTf), silver trifluoroacetate(AgOCOCF₃), silver perchlorate (AgClO₄), silver hexafluorophosphate(AgPF₆), silver sulfate (Ag₂SO₄), Nickel(II) triflate (Ni(OTf)₂),copper(II) triflate (Cu(OTf)₂), zinc triflate (Zn(OTf)₂), or aluminumethoxide (Al(OEt)₃). Conductivity changes can be monitored by includinga conductive material, such as a carbon nanotube, graphite, graphene,semiconductor nanowire, metal nanowire, or conducting polymer.Fluorescence changes can be monitored by including a luminescentmaterial, such as a conjugated molecule, a conjugated polymer, or aninorganic phosphor. The region is then monitored for color, absorbance,fluorescence, conductivity, or a combination thereof.

Methods of Quantifying the Amount of an Analyte

The amount of an analyte, such as ethylene, can be quantified based onthe color change. Alkenes of interest can be differentiated bycorrelating alkene reactivity with initial and final properties of thereaction (herein described as “reaction property profile”). Thecomposition of matter that is responsible for these reaction propertyprofiles. This disclosure describes the manner in which devices mayexploit these reaction property profiles such that a readout can be usedto detect alkenes selectively and determine cumulative device exposureto alkenes. The detection can be colorimetric, fluorescence, orconductivity.

Colorimetric systems can have certain advantages. Using standard imageprocessing software, the photographs can be used to determine the RGBvalues of the reaction mixtures. The RGB color space value can be usedas one variable to differentiate alkene and/or alkyne classes.

Euclidean distance is one variable that can be used to differentiatealkene and/or alkyne classes. In addition, Euclidean distance is onevariable that can be used to determine extent of reaction and thereforecumulative ethylene exposure to the device to quantify ethylene,preferably if starting quantities of alkene detecting material areknown.

Absorption-based concentration measurement can also be used to quantifyan analyte, such as ethylene. The absorbance of a material varieslinearly with both the cell path length and the analyte concentration.These two relationships can be combined to yield a general equation ofBeer's Law.A=εlc

A is the absorbance of the solution and is measured by a spectrometer.The quantity ε is the molar absorptivity or the extinction coefficient;l is the length of solution light passes through (cm); c isconcentration of the solution. As the reaction proceeds, theconcentration of the solution changes, which can be detected throughspectroscopic measurement.

UV-VIS spectroscopic measurements of a solution can be used to determinethe observed rate constant, k_(obs), which can then be used insubsequent calculations to determine the second-order rate constant, k₂.Each combination of compound and/or complex X and alkene and/or alkyne Yhas a unique k₂. Thus, k₂ can be one variable on which to differentiatealkene and/or alkyne classes.

A Detector for Detecting an Analyte

A detector for detecting an analyte can include a housing including adetection region comprising a compound having an extrudable group andcapable of undergoing Diels-Alder reaction with the analyte including acarbon-carbon multiple bond moiety. The housing can be a tube, solid,formulation, solid support, or solid substrates. The device can beincorporated into solid formulations or solid supports or substrates,such as paper, plastic, rubber, virtually any kind of polymer notcontaining sufficiently reactive alkenes and/or alkynes, as well asliquids such as inks.

FIG. 1 is a block diagram showing one example of a detecting device. InFIG. 1, 10 is a housing, which can be a tube, solid, formulation, solidsupport, or solid substrates, as well as liquids such as inks, and 20 isa detection region where a compound having an extrudable group canundergo a Diels-Alder reaction with the analyte including acarbon-carbon multiple bond moiety.

EXAMPLES

Selective Reaction and Detection of an Analyte

FIG. 2 is a demonstration of selective alkene detection. To Bptzdissolved in solvent was added alkene (molar excess). Experiments wereconducted under ambient atmosphere and temperature, except in the caseof C₂H₄, which was sparged through solution for 30 s at 50 SCFH and leftunder 1 atm of ethylene. Results were observed and recorded after 6.5hours of reaction. A sterically unencumbered alkene (C₂H₄) readilyreacts with Bptz. A sterically congested alkene (1,1′-diphenylethylene)does not.

Color Change after Diels-Alder Reaction

FIG. 3A shows time based color change when the solution contains silvertriflate and bptz. FIG. 3B shows one reaction example. FIG. 4 shows theextent of color change post-ethylene addition when the solution containsdifferent kinds of silver salt.

Quantify the Amount of an Analyte

FIG. 5 shows photographs of vials containingbis-2-pyridyl-1,2,4,5-tetrazine and additive in dichloromethane, beforeexposure to ethylene and after 1.5 h of exposure to a headspace of C₂H₄(1 atm) at room temperature. Using standard image processing software,the photographs can be used to determine the RGB values of the reactionmixtures. The RGB color space value can be used as one variable todifferentiate alkene and/or alkyne classes. FIG. 6 shows euclideandistance as a quantitative representation of “reaction color profile.”

In FIG. 5 and FIG. 6 , to an oven dried Teflon stir bar equipped, Teflonseptum equipped scintillation vial that was evacuated and backfilledwith argon (×1) was added metal salt (M). Each metal salt was weighedout in duplicate (one for vial containing M+Bptz and one for controlcontaining solely M). Separately, a 250-mL round bottom flask (RBF) wasequipped with a Teflon-coated stir bar and rubber septum, evacuated andbackfilled with argon, and flame dried under vacuum. To it was addedBptz (178.1 mg) and dry dichloromethane (DCM) (150.0 mL) via cannula, tofurnish a 0.05 M stock solution of Bptz (bright pink homogenoussolution). To each vial containing M was added 0.05 M Bptz solution (10mL) and each was stirred vigorously for 10 minutes, resulting in a rangeof colors (see FIG. 5 , t=0 h). Each solution was then sparged with C₂H₄for 30 s at a flow rate of 50 SCFH. The solutions were then photographedonce per minute for 2 hours.

From the photographs, the solution colors were quantified in AdobePhotoshop using the color picker function to generate RGB color spacevalues. The average of three picked values was used for each reportedvalue. The total Euclidean distance (d) traveled over the course of thereaction (net reaction color profile) was determined by equation 1:d(C _(f) ,C _(i))square root[(r _(f) −r _(i))²+(g _(f) −g _(i))²+(b _(f)−b _(i))²]  (1)

Where C stands for color; r, g, b, stand for red, green, and blue,respectively; and the subscripts _(f) and _(i) denote final and initial.

FIG. 7 shows representative example of a UV-VIS spectrum of Bptz inorganic solvent. Depicted are the three primary absorption bands thatcan be used to monitor the course of the reaction. The color perceivedby the human eye is due to the absorption band which has a λ_(max) at˜546 nm.

FIG. 8 shows graphical representation of UV-VIS spectroscopicmeasurements of λ_(max) (λ_(max)=545 nm) of a solution of Bptz inacetonitrile (MeCN) when exposed to a gross molar excess of C₂H₄ (1 atm)over time. To an ethylene purged, quartz cuvette (4 mL) equipped withsilicone septum was added Bptz in acetonitrile (MeCN) (200 uM) (2 mL).The vial was shaken rapidly, and the absorbance at 538 nm was observedat time intervals (1 s) for 130 m. Using the molar absorptivity, theabsorbance values were then converted to concentration of Bptz, [Bptz],and the plot of Ln([Bptz]) vs. t depicted was generated. Performing alinear regression on data points from t=0 to t=15 m yielded a line witha slope that is k_(obs). k_(obs) can then be used in subsequentcalculations to determine the second-order rate constant, k₂. Eachcombination of compound and/or complex X and alkene and/or alkyne Y hasa unique k₂. Thus, k₂ can be one variable on which to differentiatealkene and/or alkyne classes.

FIG. 9 shows kinetics experiment results in Nitromethane (MeNO₂),Dimethylformamide (DMF), and MeCN. Cuvette (4 mL) equipped with siliconeseptum was purged with C₂H₄; bptz in solvent (2 mL; 200 μM) was injectedinto cuvette equipped with bubbler; cuvette was shaken up and downrapidly; measurement then began immediately at λ_(max) (nm).

Color Based Detectors for Detecting an Analyte

FIG. 10 shows photographs of alkene sensing material incorporated into aglass tube containing glass wool (glass wool surrounding thepink-colored alkene sensing material). The depicted glass tube (device)is equipped with Leur-type syringe tips, enabling connection togas-stream inlet and outlet tubing. The device was exposed to acontinuous stream of 1,000 ppm C₂H₄ in N₂ carrier gas at a flow rate of0.5 L/min. An indicating amount of Bptz impregnated polystyrene fiberswas added to the glass tube containing glass wool. These fibers were theby-product of the fabrication process described in the experimentalpertaining to FIG. 11 . The tube was capped with modified plasticsyringe tips, and was equipped to a KinTech gas mixer. The tube was thensubjected to 1,000 ppm C₂H₄ in N₂ at a flow rate of 0.5 L/min.Photographs were taken at t=0 h, 24 h, and 48 h to monitor thedecolorization process.

FIG. 11 shows photographs of Bptz impregnated polystyrene coatingcovering a printed QR code on copy paper. FIG. 11 (panel A) shows aphotograph of QR code printed in pink on copy paper. Before exposure toC₂H₄, the coating is pink and the QR code is unreadable, as shown inFIG. 11 (panel B). After exposure to C₂H₄ (1 atm) for 1 h, thepolystyrene coating has decolorized, rendering the QR code readable bymachines (e.g. personal hand-held devices such as camera phones), asshown in FIG. 11 (panel C). To a solution of Bptz in DCM (0.01 M) wasadded polystyrene such that the final volume ratio was 4:1 solution:polystyrene (v/v), resulting in a pink, viscous, homogenous solution.Separately, a piece of copy paper with printed QR code had been affixedwith double sided tape to a glass cover slide and fitted to aspin-coater. The copy paper sample was spun at 2,000 rpm and the stocksolution described above was expelled from a syringe directly onto itfrom above, at a rate of ˜1 mL/min. After removing any remaining DCMunder vacuum, the resulting plastic coated QR code was exposed to C₂H₄(1 atm) in an air-tight glass container for 2 hours. Before and afterphotographs were taken to document the color change.

FIG. 12 shows photographs of various standardized, machine-readablerepresentations of data printed using inks created with Bptz as pigment:UPC code in FIG. 12 (panel A), Quick Read (QR) code in FIG. 12 (panelB), USPS data matrix in FIG. 12 (panel C), and GS1 DataBar in FIG. 12(panel D). FIG. 12 (panel E) shows a photograph depicting Bptz ink (pinkcolored solution indicated by arrow) with an example of a GS1 DataBarthat it was used to print on copy paper.

FIG. 13 (panel A-1) and FIG. 13 (panel A-2) shows photographs ofstandardized, machine-readable representations of data printed usinginks created with Bptz as pigment before exposure to C₂H₄; FIG. 13(panel B-1) and FIG. 13 (panel B-2) shows photographs of therepresentations after exposure to C₂H₄ (1 atm) for 48 h.

Accompanying FIG. 12 and FIG. 13 , an empty HP desktop inkjet printercartridge was charged with a solution of Bptz in 2:1 (v/v) H₂O: Acetone(0.01 M). It was quickly loaded into an HP inkjet printer, and thedepicted images were printed in greyscale from a .PDF file, resulting inlight-pink images on copy paper. The images were made darker by printingthe same image multiple times (5-10 passes). The tetrazine-printedimages (i.e. codified information) were exposed to C₂H₄ (1 atm) in asealed glass container for 2 d, resulting in a color change from pink tofaint yellow.

Effect of Acids

To an ethylene purged quartz cuvette equipped with silicone septum wasadded a solution of Bptz+trifluoroacetic acid (TFA) (200 uM in Bptz) intoluene (2 mL). Two and one-half molar equivalents of TFA wereevaluated, and the growth of the UV-VIS absorption band at 410 nm wasobserved over time, to yield insight into the dependence of k_(obs) onthe presence of Bronsted acid. FIG. 14 is a graph showing a rateenhancement when Bptz is in the presence of Bronsted acid, in this caseTFA. It shows that this rate enhancement is dependent on the molarequivalents of TFA present.

In another example, a quartz cuvette containing a solution ofBptz+AgOCOCF₃ (1:1; 0.5 mM in Bptz) was charged with 1-hexene. UV-VISmeasurements were started immediately, and were recorded at 0.1 mintervals for 15 m. FIG. 15 shows the change in UV-VIS spectra of 1:1bptz:AgOCOCF₃ over a 15 minute time course. It shows that a new, strongabsorption band forms in the visible (˜420 nm) which has a strong molarabsorptivity. Thus, using silver salts allows us to monitor twovariables simultaneously: disappearance of absorption band at 545 nm,and appearance of a new, stronger band at 420 nm. Note also that thereis a rate enhancement associated with the silver salt, as this reactionwas done in 15 minutes as opposed to the usual 1 h for naked Bptz. Ingeneral, Lewis acids can be rate-enhancing additives.

Fabrication of Selective Alkene Chemiresistors from NanostructuredCarbon and Alkene Selector Molecules

Evaporation of Gold on Paper.

Gold (Au) electrodes were deposited on paper. See, for example, Mirica,K. A. et al., Proc. Natl. Acad. Sci. 2013, E3265-E3270, which isincorporated by reference in its entirety.

Stock Solution.

To a scintillation vial (20-mL) was added pristine single-walled carbonnanotubes (SWCNTs) (3.98 mg) and ortho-dichlorobenzene (o-DCB). The vialwas capped with a plastic screw-cap, and sonicated in a water bath atroom temperature for 10 minutes, resulting in a black heterogenoussolution. To a separate scintillation vial (20-mL) was added alkeneselector molecule (e.g. tetrazine) (S) (2.02 mg), o-DCB (1.5 mL) andchloroform (CHCl₃) (0.5 mL). The vial was capped with a plasticscrew-cap and sonicated in a water bath at room temperature for 10minutes, resulting in a colored homogenous solution (color dependent onS). The above solutions were combined by way of a plastic syringe orglass pipette and mixed thoroughly with a magnetic stir bar or byinverting (×10) to form a black heterogenous stock solution.

Fabrication of an Array of Sensors.

Sensors were fabricated by dropcasting the freshly made stock solutionwith a capillary tube (5 μL) in between gold electrodes on a substrate(weigh paper or glass). The solvent was removed by placing theelectrodes into an evacuated chamber. After removal of the solvent,residual material was visibly adhered onto the surface of the substrate,bridging the gold electrodes. The resistance of the chemiresistor, R,was measured using a multimeter. The above process was repeated until1.0 kΩ<R<20.0 kΩ for each sensor.

Sensing Measurements.

The array of sensors was mounted onto a glass slide (25 mm×75 mm×1 mm)with double-sided Scotch tape. The array was then inserted into a 2×30pin edge connector, which made electrical contact with each of the goldelectrodes within the array. The edge connector was then connected tothe potentiostat via a breadboard (DigiKey). For sensing measurements,the devices were enclosed within a custom-made gas-tight Teflon chambercontaining an inlet and outlet port for gas flow. The inlet port wasconnected to a gas delivery system, and the outlet port was connected toan exhaust vent. Measurements of conductance were performed under aconstant applied voltage of 0.1 V using PalmSense EmStat-MUX equippedwith a 16-channel multiplexer (Palm Instruments BV, The Netherlands).Data acquisition was done using PSTrace 2.4 software provided by PalmInstruments.

Delivery of Ethylene.

Delivery of controlled concentrations of ethylene to devices was carriedout using Smart-Trak Series 100 gas mixing system at total flow ratesbetween 2.0 mL/min and 1.0 L/min.

FIG. 16 shows test results of sensors deposited using drop-cast(o-DCB/CHCl₃), with weigh paper as substrate, and Au (120 nm) aselectrodes. SWCNT, ultrapure, was obtained from NanoC, and bptz, 96%,was obtained from Aldrich. Sensor composition 1:10 Tz/C (mol/mol) wasused. Tz is generic for any tetrazine, such as Bptz or Tz-2. C is “molesof carbon atoms in SWCNTs” and is the generic case. FIG. 16 shows thatSWCNT-bptz mixtures become less conductive in the presence of ethylene.In FIG. 16 , a device fabricated one day prior to use (Group A, greentrace) with a Bptz/SWCNT (2:1 Bptz/SWCNT (wt/wt)) chemiresistordisplayed a dosimetric response toward pure ethylene (1 atm), with a 50%change in conductivity with respect to it's original state. Similarly,three devices fabricated four days prior to use with Bptz/SWCNTchemiresistors ((2:1 Bptz/SWCNT (wt/wt)) displayed dosimetric responsestoward pure ethylene (1 atm), with a 35% change in conductivity withrespect to it's original condition for one device (Group A; red trace),and a 30% change in conductivity with respect to their originalconditions (Group A; yellow and blue traces) for the other two devices.Three devices fabricated one day prior to use with Tz-2/SWCNTchemiresistors (2:1 Tz-2/SWCNT (wt/wt)) displayed dosimetric responsestoward pure ethylene (1 atm), with a 15% change in conductivity withrespect to their original conditions (Group B; red, yellow, and bluetraces). Three devices fabricated with pristine SWCNTs four days priorto use were employed as a negative control (Group C; red, yellow, andblue traces). They did not display a dosimetric response; rather, theydisplayed a reversible response that was less than 3% change inconductivity relative to their original conditions.

FIG. 17 is a demonstration of selective electron-rich alkene detection.To Bptz dissolved in chloroform (5 mM) was added alkene (molar excess).Experiments were conducted under ambient atmosphere and temperature.Results were observed and recorded with a digital camera after 0, 10,30, 60, and 90 minutes of reaction. Electron-rich alkenes (1-hexene,isoprene, styrene) reacted readily with Bptz. Electron-poor alkenes(acrylonitrile, ethyl acrylate) reacted slowly or not at all.

FIG. 18 is a demonstration of selective electron-poor alkene detection.To 1,3-diphenylisobenzofuran (1 mM) dissolved in chloroform was addedalkene (molar excess). Experiments were conducted under ambientatmosphere and temperature. Results were observed and recorded with adigital camera after 0, 10, 30, 60, and 90 minutes of reaction.Electron-poor alkenes (acrylonitrile, ethyl acrylate) reacted readilywith 1,3-diphenylisobenzofuran. Electron-rich alkenes (1-hexene,isoprene, styrene) reacted slowly or not at all.

FIG. 19 is a demonstration of selective electron-poor alkene detection.To tetraphenylcyclopentadienone (tetracyclone) (1 mM) dissolved inchloroform was added alkene (molar excess). Experiments were conductedunder ambient atmosphere and temperature. Results were observed andrecorded with a digital camera after 0, 1, and 18 hours of reaction.Electron-poor alkenes (acrylonitrile, ethyl acrylate) reacted after 18hours with tetracyclone. Electron-rich alkenes (1-hexene, isoprene,styrene) reacted slowly or not at all.

FIG. 20 is a demonstration of other tetrazines selectively reaction withelectron rich alkenes. To tetrazines (Tz-2-Tz-6) (1 mM) dissolved inchloroform was added alkene (molar excess). Experiments were conductedunder ambient atmosphere and temperature. Results were observed andrecorded with a digital camera after 0 and 60 minutes of reaction. Alltested alkenes reacted with Tz-3, although acrylonitrile reacted theslowest. Electron-rich alkenes (1-hexene, isoprene, styrene) reactedwith slowly with Tz-4. Electron-poor alkenes (acrylonitrile, ethylacrylate) reacted very slowly or not at all with Tz-4. Electron-richalkenes (1-hexene, isoprene, styrene) reacted slowly with Tz-2.Electron-poor alkenes (acrylonitrile, ethyl acrylate) reacted veryslowly or not at all with Tz-2. Electron-rich alkenes (1-hexene,isoprene, styrene) reacted slowly with Tz-5. Electron-poor alkenes(acrylonitrile, ethyl acrylate) reacted very slowly or not at all withTz-5. Electron-rich alkenes (1-hexene, isoprene, styrene) reacted slowlywith Tz-6. Electron-poor alkenes (acrylonitrile, ethyl acrylate) reactedvery slowly or not at all with Tz-6.

FIG. 21 is a demonstration of complementary reactivity of Bptz and1,3-diphenylisobenzofuran. To Bptz dissolved in chloroform (5 mM) wasadded alkene (molar excess). To 1,3-diphenylisobenzofuran (1 mM)dissolved in chloroform was added alkene (molar excess). Experimentswere conducted under ambient atmosphere and temperature. Results wereobserved and recorded with a digital camera after 0 and 30 minutes ofreaction. Electron-poor alkenes (acrylonitrile, ethyl acrylate) reactedreadily with 1,3-diphenylisobenzofuran with a concomitant loss of color.Electron-rich alkenes (1-hexene, isoprene, styrene) reacted slowly ornot at all with 1,3-diphenylisobenzofuran without a perceivable loss ofcolor. Electron-rich alkenes (1-hexene, isoprene, styrene) reactedreadily with Bptz with a concomitant change in color. Electron-pooralkenes (acrylonitrile, ethyl acrylate) reacted slowly or not at allwith Bptz with little or no perceivable change in color.

Other embodiments are within the scope of the following claims.

What is claimed is:
 1. A method of detecting an analyte in a sampleincluding a carbon-carbon multiple bond moiety comprising: exposing adetection region of a detector including a compound having an extrudablegroup and capable of undergoing Diels-Alder reaction with the analyteincluding a carbon-carbon multiple bond moiety to the sample, anddetecting a property change of a reaction mixture comprising theheteroaromatic compound based on the presence of the analyte in thesample wherein the reaction mixture includes a copper salt, a nickelsalt, a silver salt, a zinc salt, an aluminum salt, or a gold salt. 2.The method of claim 1, wherein the analyte is ethylene.
 3. The method ofclaim 1, wherein detecting the property change includes monitoringabsorbance of the detection region, fluorescence of the detectionregion, or conductivity of the detection region.
 4. The method of claim1, further comprising determining a reaction rate constant of theanalyte with the compound in the detection region of the detector. 5.The method of claim 4, further comprising quantifying the amount of theanalyte using the rate constant.
 6. The method of claim 1, furthercomprising differentiating alkene classes, alkyne classes, or alkene andalkyne classes by correlating reactivity with initial and final colorsof the reaction mixture.
 7. The method of claim 1, further comprisingquantifying the amount of the analyte using Red Green Blue colormapping.
 8. The method of claim 1, further comprising differentiatingalkene classes, alkyne classes, or alkene and alkyne classes using RedGreen Blue color space value.
 9. The method of claim 1, furthercomprising quantifying the amount of the analyte using Euclideandistance determined from the initial and final colors of the reactionmixture.
 10. The method of claim 1, further comprising differentiatingalkene classes, alkyne classes, or alkene and alkyne classes usingEuclidean distance.
 11. The method of claim 1, further comprisingreading a machine-readable pattern in the detection region when thepattern appears or reading a machine-readable pattern in the detectionregion when the pattern vanishes.
 12. The method of claim 1, wherein thereaction mixture is in a solvent, a solid formulation or a solidsubstrate.